HOT FIREPol® Probe qPCR Mix Plus (Capillary)

A 5x concentrated, ready-to-use master mix optimized for real-time quantitative PCR assays and supplied by Solis BioDyne. It contains all the components necessary to perform qPCR, with the exception of template, primers, and probe. The formulation includes optimized components together with HOT FIREPol® DNA Polymerase in a proprietary reaction buffer designed to enable detection of low copy number targets, and is optimized for DNA hydrolysis probes, exploiting 5' flap endonuclease activity. HOT FIREPol® DNA Polymerase is activated by a 12 minute incubation step at 95°C, which prevents extension of non-specifically annealed primers and primer-dimer formation at low temperatures during reaction setup.

Recommended applications include detection and quantification of DNA and cDNA targets, gene expression profiling, microbial detection, and viral load determination. The mix composition comprises HOT FIREPol® DNA Polymerase, a 5x Probe qPCR buffer (providing 3 mM MgCl2 in the 1x solution, from a 15 mM MgCl2 stock), dNTPs, and BSA. A typical 20 µl reaction uses 4 µl of the 5x mix with 200–400 nM of each primer and 100–250 nM probe. Recommended template concentrations are 0.1 pg/µl–10 ng/µl for cDNA and 10 pg/µl–4 ng/µl for gDNA. The suggested cycling protocol is an initial activation of 12 min at 95°C, followed by 40 cycles of 15–20 s at 95°C (denaturation) and 60 s at 60°C (annealing/extension), with the annealing temperature generally set 2–5°C below the primer melting temperature. Reaction setup is recommended at room temperature, ideally under a laminar flow hood or in a PCR box to prevent contamination.

Routine storage is at -28°C to -18°C, and shipping or temporary storage for up to 1 month at room temperature has no detrimental effects on the quality of the product. The product is manufactured under an ISO 9001 and ISO 13485 certified quality management system and is supplied for research use only. FIREPol® is an EU registered trademark of Solis BioDyne OÜ. Users are advised to wear suitable protective clothing, gloves, and safety glasses when handling the product and its components.